Exosome in Methamphetamine and HIV-associated Neurodegeneration Abstract Exosomes are a class of extracellular vesicles that have been indicated as emerging mediators of cell-to- cell communication. Viral molecules present in exosomes derived from HIV-infected cells have been implicated as critical transmitters of intercellular viral spread, representing a receptor-independent mode of infection (Trojan exosome hypothesis). However, whether exosomes from HIV-infected cells are able to cross blood-brain barrier (BBB) and target brain viral reservoirs, particularly in the context of psychostimulant drugs, remains unclear. Our early studies showed that methamphetamine (METH) could modulate the expression of a variety of HIV intracellular restriction factors, thus compromising host cell innate immunity. Recently, we revealed that exosomes released from immune-primed brain microvascular endothelial cells (BMVEC) are able to incorporate, transfer functional RNA/proteins to macrophages and suppress HIV replication. Based on these earlier studies, we hypothesize that METH can compromise the innate immunity of BMVEC, thus affecting exosome-mediated horizontal transmission of immunoregulatory activity from BBB to monocytes, facilitating HIV dissemination to brain and resulting in neurodegeneration. We propose three specific aims to address these hypotheses: Aim 1. To determine the effect of METH on exosomal miRNA profile of BMVEC and how this modulation affects monocyte activation and susceptibility to HIV infection; Aim 2. To examine the role of exosomes derived from HIV-infected macrophages on BBB integrity and neurodegeneration in the context of METH use; Aim 3. To characterize plasma exosomal miRNA signatures of METH users with or without HIV infection and validate their functional significance in central nervous system (CNS) homeostatic regulation. Data arising from this study with both in vitro and in vivo systems will be clinically relevant and important to our understanding of the role of exosomes in brain homeostasis. It is also significant in terms of developing plasma exosomal miRNA signature as a biomarker for METH- and/or HIV-associated neurodegeneration.